Nanoparticle targeting of cathepsin-L inhibitor and doxorubicin in breast cancer

描述（由申请方提供）：对化疗产生耐药性代表肿瘤细胞的适应性生物学反应，导致治疗失败和患者复发。如果治疗要成功根除肿瘤，迫切需要克服这个问题。肿瘤细胞不可逆的生长停滞（衰老）是对暴露于化学治疗剂的应激的早期细胞反应。那些能够绕过衰老的细胞最终表现出对化疗的抵抗力。最近的报道为研究组织蛋白酶L在化疗药物耐药过程中的作用提供了一个有说服力的理论基础。我们发现，使用化学抑制剂或siRNA特异性靶向组织蛋白酶L，迫使癌细胞进入不可逆的生长停滞状态，并抑制对药物的耐药性。我们还证明了组织蛋白酶L抑制剂逆转体内耐药性的能力，利用荷阿霉素耐药神经母细胞瘤细胞系SKN-SH/R耐药肿瘤的异种移植物的裸鼠。因为许多化疗药物显示全身毒性，并且因为这些研究中使用的组织蛋白酶L抑制剂是易生物降解的肽，我们将通过使用靶向肿瘤新血管形成部位（1v整联蛋白）或通过HER 2抗体靶向MCF 7肿瘤细胞的PEG-PLGA纳米颗粒包封药物来解决这些限制。雌性裸鼠将具有耐药性MCF 7人乳腺癌细胞原位植入第四乳腺中。我们将研究两种市售组织蛋白酶L抑制剂Z-Phe-Try（t-Bu）-重氮甲基酮和1-萘磺酰基-Ile-Trp-醛单独给药或与多柔比星联合给药对肿瘤细胞生长和耐药性的影响。具体目标1：在裸鼠中进行纳米颗粒靶向治疗研究之前，将在肿瘤血管生成和生长的鸡胚绒毛尿囊膜（CAM）肿瘤植入模型中进行体内初步实验以确定纳米颗粒的最佳制剂。该模型允许在体内预筛选生物活性，同时限制使用更敏感和昂贵的鼠物种。具体目标二：将在小鼠乳腺癌模型中测试在CAM模型中显示最佳抗肿瘤和抗血管生成活性的纳米颗粒制剂。在这些研究中，我们将评估组织蛋白酶L抑制剂在降低阿霉素耐药性方面的有效性，这一点通过改善抗肿瘤活性得到证实。将比较非靶向和靶向治疗的抗肿瘤疗效和多柔比星相关毒性。癌细胞具有对化疗药物产生耐药性的独特能力，因此研究逆转这种现象的方法对癌症患者的治疗具有重要价值。该项目将在乳腺癌小鼠模型中使用两种药物的组合，其中一种药物会削弱癌细胞产生耐药性的能力。一种新的技术，使用纳米粒子封装测试药物，将进行测试，以确定这些纳米粒子是否可以改善药物的输送和最大限度地减少相关的毒性。

DESCRIPTION (provided by applicant): The development of resistance to chemotherapy represents an adaptive biological response by tumor cells that leads to treatment failure and patient relapse. There is an urgent need to overcome this problem if treatments are to be successful in eradicating tumors. Tumor cell irreversible growth arrest (senescence) is an early cellular response to the stress of exposure to chemotherapeutic agents. Those cells that are able to bypass senescence ultimately exhibit resistance to chemotherapy. Recent reports provide a persuasive rationale for studying the role of cathepsin L in the process of chemotherapeutic drug resistance. We showed that specific targeting of cathepsin L using chemical inhibitors or siRNA to this molecule forced cancer cells into a state of irreversible growth arrest and suppressed development of resistance to drugs. We also demonstrated the ability of cathepsin L inhibitor to reverse drug resistance in vivo utilizing nude mice bearing xenografts of doxorubicin resistant neuroblastoma cell line SKN-SH/R drug resistant tumors. Because many chemotherapeutic agents show systemic toxicities, and because cathepsin L inhibitors to be used in these studies are peptides subject to biodegradation, we will address these limitations by encapsulation of agents using PEG-PLGA nanoparticles targeted to either the sites of tumor neovascularization (1v integrin) or to MCF7 tumor cells via HER2 antibody. Female nude mice will have drug-resistant MCF7 human breast cancer cells implanted orthotopically into the fourth mammary gland. We will study two commercially available Cathepsin L inhibitors, Z-Phe-Try (t-Bu)-diazomethylketone and 1-Naphthalenesulfonyl-Ile-Trp- Aldehyde given either alone or with doxorubicin with respect to their effects on tumor cell growth and drug-resistance. Specific Aim 1: Prior to performance of nanoparticle-targeted therapy studies in nude mice, preliminary experiments to determine optimum formulations of nanoparticles will be performed in vivo in the chick chorioallantoic membrane (CAM) tumor implant model of tumor angiogenesis and growth. This model permits in vivo pre-screening for bioactivity while limiting the use of more sentient and costly murine species. Specific Aim 2: Nanoparticle formulations that show optimum anti-tumor and anti-angiogenesis activity in the CAM model will be tested in the mouse breast cancer model. In these studies, we will evaluate the effectiveness of cathepsin L inhibitors in reducing doxorubicin resistance as evidenced by improved anti-tumor activities. Comparisons will be made between non-targeted and targeted therapies with respect to anti-tumor efficacy and doxorubicin-associated toxicities. Cancer cells have the unique ability to develop resistance to chemotherapeutic drugs, and so research on ways to reverse this phenomenon would have significant value in the treatment of cancer patients. This project will use a combination of two drugs, one of which impairs the cancer cell's ability to develop drug resistance, in a mouse model of breast cancer. A novel technology, the use of nanoparticles to encapsulate the test drugs, will be tested to determine whether these nanoparticles can improve the delivery of drugs and minimize the associated toxicities.